The purpose of this study was to understand the culture, values, skills and activities of staff involved in education and prevention activities in community-based AIDS Service Organizations (ASOs) in Ontario, Canada, and to understand the role of evaluation research in their prevention programming. In this qualitative study, 33 staff members from 11 ASOs participated in semi-structured interviews that were analyzed using the grounded theory approach. ASO staff experience tension between a historical grassroots organizational culture characterized by responsiveness and relevance and a more recent culture of professionalization. Target populations have changed from being primarily gay men to an almost unlimited variety of communities. Program emphasis has shifted from education and knowledge dissemination to a broadly based mandate of health promotion, community development, and harm reduction. Integration of evidence of effectiveness, social-behavioral theory, or systematic evaluation is uncommon. Understanding these points of tension is important for the nursing profession when it is engaged with ASOs in programming or evaluation research.